Biochimica et Biophysica Acta, 708 (1982) 225-232 225 Elsevier Biomedical Press BBA 31339 SUSCEPTIBILITY TO PROTEINASES OF YEAST ENZYMES SELECTIVELY MODIFIED BY FATIN ACIDS N. BURLINI a, p. TORTORA a, G.M. HANOZET a, M.T. VINCENZINI b, p. VANNI b and A. GUERRITORE a a Department of General Physiology and Biochemistry, University of Milan, Via Saldini 50, Milan, and b Department of Biochemistry, University of Florence, Viale Morgagni 50, Florence (Italy) (Received June 14th, 1982) Key words: Enzyme modification," Proteinase susceptibility; Fatty acid," Glucose-6-phosphate dehydrogenase; Phosphoglycerate kinase," A Icohol dehydrogenase; (Yeast) To investigate a possible correlation between selective modification and degradation of enzymes, the susceptibility to intracellular yeast proteinases A and B of yeast enzymes treated with fatty acids was tested. Enzymes used were glucose-6-phosphate dehydrogenase (EC 1.1.1.49) and 3-phosphoglycerate kinase (EC 2.7.2.3), which are sensitive to the denaturing modification caused by fatty acids, and alcohol dehydrogenase (EC 1.1.1.1) which is insensitive. Proteinases and substrate enzymes were all pure preparations. Without modification by fatty acids, at neutral pH, the three enzymes are remarkably resistant to degradation by both proteinases. Treatment with myristic or oleic acid definitely enhances the susceptibility to proteolysis of the sensitive glucose-6-phosphate dehydrogenase and 3-phosphoglycerate kinase, whereas it leaves negligible that of the insensitive alcohol dehydrogenase. The selective effect of fatty acids on the degradation is pH-depen- dent: with proteinase A it was lost at acidic pH. Since intracellular levels of free fatty acids near or even higher than 1 mM were actually measured in yeast cells, it is possible that free fatty acids, in some cellular conditions, affect yeast enzyme composition. However, the control of specific enzyme degradation in yeast is still an open question. Introduction A plausible model of intracellular enzyme de- gradation includes the conversion of the holo- form into the apo- form, possible further modifica- tion by interaction with cellular agents, and then action on the modified protein of proteinases, either free in the cytosol or bound in lysosome or vacuole membranes. The basis of selectivity could be the substrate protein or the degrading system. When protein modification is the rate-limiting step, the change in hydrophobicity of the molecular surface is one of the important determinants of the process [ 1,2]. The interaction between enzymes and free fatty acids represents a process of denaturing modifica- 016%4838/82/0000-0000/$02.75 © 1982 Elsevier Biomedical Press tion, which can change the hydrophobicity of the protein surface [3]. Denaturation of yeast enzymes by fatty acids was found to be selective [4] and partially reversible [3]. The data reported in the present paper relate the modification by fatty acids of some yeast enzymes to their susceptibility to purified yeast proteinases, at concentrations of free fatty acids that correspond to the levels actu- ally found in the yeast cell. Enzymes tested were glucose-6-phosphate dehydrogenase (EC 1.1.1.49) and 3-phosphoglycerate kinase (EC 2.7.2.3), which are sensitive to long-chain fatty acids; and alcohol dehydrogenase (EC 1.1.1.1), which is insensitive [4]. Yeast proteinases were intracellular proteinase A (EC 3.4.23.6.) and B (EC 3.4.22.9).